CbrAB-dependent regulation of pcnB, a poly(A) polymerase gene involved in polyadenylation of RNA in Pseudomonas fluorescens

Abstract

CbrB is a global σ 54-dependent regulator required for nutrient acquisition in Pseudomonas. Located downstream of cbrB on the Pseudomonas fluorescens SBW25 chromosome is pcnB, a putative poly(A) polymerase gene. Presence of a σ 54 promoter in the intergenic region of cbrB and pcnB led to the hypothesis that CbrB regulates pcnB expression in a σ 54- dependent manner. Here we show that transcription of pcnB is CbrB dependent. However, 5′-RACE analysis of the pcnB transcript using primers located in the pcnB coding region shows that transcription starts immediately upstream of the putative ATG site at a σ 70-like promoter. Deletion of pcnB caused ~80% decrease of ployadenylated 23S rRNA; growth of the pcnB mutant was compromised in a range of labora torymedia and on sugar beet seedlings. Further 5′-RACE analysis confirmed the existence of the predi cted σ 54 promoter. Genetic analysis showed that the σ 54 promoter drives expression of crcZ, a homologue of the recently described small RNA from Pseudomonas aeruginosa, in a CbrB-dependent manner. Taken together, our data show that both pcnB and crcZ are part of the CbrB regulon. Moreover, the data draw further attention to the central regulatory role of CbrB and provides a link between mRNA degradation and cellular catabolism. © 2010 Society for Applied Microbiology and Blackwell Publishing Ltd.